In International Patent Application No. PCT/GB87/00365 (International Publication No. WO 87/07295) enzyme electrodes capable of responding amperometrically to the catalytic activity of the enzyme in the presence of its respective substrate are described comprising an enzyme immobilised or adsorbed onto the surface of an electrically conductive support member which consists of or comprises a porous layer of resin-bonded carbon or graphite particles, said particles having intimately mixed therewith, or deposited or adsorbed onto the surface of the individual particles prior to bonding to form said layer, a finely divided platinum group metal, thereby to form a porous, substrate layer onto which said enzyme is adsorbed or immobilised and comprising a substantially homogeneous layer of resin-bonded carbon or graphite particles, with said platinum group metal dispersed substantially uniformly throughout said layer. The preferred substrate materials are resin bonded platinised carbon paper electrodes comprising platinised carbon powder particles having colloidal platinum adsorbed on the surface of the particles and bonded onto a carbon paper substrate using a synthetic resin, preferably polytetrafluoroethylene, as the binder. The preferred enzyme electrodes are glucose oxidase electrodes comprising glucose oxidase adsorbed or immobilised onto the surface of the substrate.
In International Application No. PCT/GB88/00868 (International Publication No. WO 89/03871) similar enzyme electrodes are disclosed but using the oxides, e.g. PtO, rather than an elemental platinum group metal preadsorbed onto the resin-bonded carbon or graphite particles.
As described therein, the preferred substrate materials for those enzyme electrodes are resin bonded, platinised (or platinum oxide containing) carbon materials used hitherto as gas diffusion electrodes in fuel cells and available commercially from The Prototech Company, Newton Highlands, United States of America. Essentially such materials contain as the resin binder a relatively high melting point hydrophobic fluorocarbon resin, preferably polytetrafluoroethylene.
The manufacture of such resin-bonded platinised carbon, gas diffusion electrode materials is described in U.S. Pat. Nos. 4,044,193; 4,166,143; 4,293,396 and 4,478,696. Alternative, but similar gas diffusion electrode materials, equally suitable in accordance with the teachings of International Patent Publication WO 87/07295 as the electrically conductive substrate materials for enzyme electrodes are also disclosed in U.S. Pat. No. 4,229,490. Broadly speaking, such electrode materials are manufactured by depositing colloidal size particles of platinum or palladium, or other platinum group metal, onto finely divided particles of carbon or graphite, blending the platinised or palladised carbon or graphite particles with a fluorocarbon resin, preferably polytetrafluoroethylene, and moulding the mixture onto an electrically conductive support, preferably an electrically conductive carbon paper, or onto a filamentous carbon fibre web. Similar procedures apply to the oxide containing materials.
For use as an enzyme electrode in accordance with the teachings of International Publications WO 87/07295 and WO 89/03871, the appropriate enzyme, e.g. glucose oxidase, or mixture of enzymes, is immobilised or adsorbed onto the surface of a preformed web comprising a porous surface layer of resin-bonded platinised or palladised (those terms hereinafter being used in a generic sense to include the corresponding oxides as well as the elemental platinum group metals, unless the context requires otherwise) carbon or graphite particles, as described above. As indicated, the immobilised enzyme may simply be adsorbed onto the surface of the porous layer of resin-bonded, platinised or palladised carbon particles, or it may be covalently coupled thereto, for example, using well established enzyme immobilisation techniques, such as, for example, covalent bonding with a carbodiimide or carbonyldiimidazole reagent, covalent bonding with 1,6-dinitro-3,4-difluorobenzene, or by cross-linking with glutaraldehyde. In all cases, the enzyme or enzyme mixture is immobilised or adsorbed onto a preformed electrically conductive substrate comprising a porous layer of resin bonded platinised or palladised carbon or graphite particles moulded onto an electrically conductive substrate by the application of heat and pressure. In place of the finely divided platinum group metal, the corresponding oxides may be used, e.g. platinum or palladium oxide.
Amongst other relevant background prior art to be acknowledged herein are:
Ianiello et al (1982) Analyt. Chem. 54, 1098-1101 which describes mediatorless sensors in which glucose oxidase and L-amino acid oxidase are covalently bonded to a graphite electrode by the cyanuric chloride method;
Matsushita Electric Appliance Industry Company, Unexamined Japanese Patent Publication No. 56-16447 which discloses an enzyme electrode comprising an electrically conductive base of moulded graphite containing up to 10 parts by weight of a fluorocarbon resin binder, e.g. polytetrafluoroethylene, as the binder, and onto which is deposited by vapour phase deposition or electrolytically, a thin (less than 1 .mu.m) film of platinum. The enzyme, e.g. glucose oxidase is immobilised onto the platinised surface of the electrically conductive base, the invention allegedly overcoming the problems of immobilising an enzyme directly onto platinum; and
Matsushita Electrical Industrial Co. Ltd. (Nakamura et al) U.S. Pat. No. 4,392,933 in which is disclosed an immobilised enzyme electrode comprising an oxido-reductase enzyme, e.g. glucose oxidase, and a metal oxide, e.g. ruthenium oxide, capable of entering into the redox reaction coupled to the enzyme, the enzyme and the metal oxide, the metal oxide itself either itself forming an electron collector and conductor, or being incorporated into an electron collector and conductor material, such as graphite. When using graphite as the electron collector and conductor, the reactive metal oxide, e.g. RuO.sub.2 in powder form and graphite powder are press moulded into a powder compact or disc, and onto which the enzyme, e.g. glucose oxidase, is immobilised by cross-linking to the graphite surface, for example, with glutaraldehyde.